Use of alkylmonoglucosides as molecular vectors

ABSTRACT

The invention concerns the use as molecular vector in pharnaceutical, cosmetic and food products of alkylmonoglucosides of general formula (I) in which R1 is a linear or branched C2-C18 alkyl radical.

The present invention concerns the use of alkylmonoglucosides and moreparticularly n-butyl-α-D-monoglucopyranose, named hereafterα-butylglucoside as molecular vector, and the preparation of newcompounds obtained by grafting α-butylglucoside onto certain compoundsand their uses.

We have noted that a need existed to modify numerous cosmetic orpharmaceutical active ingredients and/or food ingredients in order toimprove:

their bioavailability

their toxicity

their liposolubility

their hydrosolubility

The present invention proposes to respond to these needs. It concernsthe use, in pharmaceutical, dermatological, cosmetic or food domains, ofalkylmonoglucosides as transcutaneous or transmucous membrane molecularvectors having the general formula:

in which R₁ is a C₂ to C₁₈ alkyl, linear or branched radical.

By “molecular vector” a compound is meant, which after chemical reactionwith an active compound gives a vectorized active compound whichpenetrates the skin or a mucous membrane more easily than the initialactive compound.

The invention also concerns the use, in a pharmaceutical,dermatological, cosmetic or food composition, of vectorized activeingredients with the general formula:

where R₂ is a —CO—R group where R is a hydrocarbonated, linear,branched, saturated or unsaturated ethylene radical; a group derivedfrom retinol (vitamin A) or from one of its derivatives, notablyretinoic acid, ascorbic acid (vitamin C) or one of its derivatives, of atocopherol amongst which are vitamin E and the D vitamins; a groupderived from polyphenols, for example a residue of polyhydroxylatedderivatives of flavan; or a radical

where X is an aliphatic chain which is functionalized or not.

In these vectorized active ingredients, the active molecule is linked bycovalence to position 6 of the vector. The invention stems from the factthat we have reduced the toxicity of exfoliant agents used in cosmeticsand/or dermatology by grafting onto α-butylglucoside.

Cosmetic and/or dermatological compositions have, amongst others, avocation of acting on the protection function of the skin whichnecessitates the direct influence of the condition of the corneal layer.It is known that if this corneal layer contains too many dead cells, itdoesn't protect any more. It must then be removed to allow another layerof cells to maintain an efficient barrier against external aggressions,and the cosmetic and dermatological active ingredients to penetrate it.Such is the known and allotted role of Alpha Hydroxy Acids or AHA.

The AHA are organic acids with an alcohol function on the neighbouringcarbon of the one (in alpha position) carrying the carboxylic acidfunction. We group together more particularly glycolic acid, lacticacid, malic acid, tartaric acid, citric acid, gluconic acid as well ascertain analogues of AHA like salicylic acid and serine in this familyof compounds.

These AHA have recognised and measurable efficacy but they also presentsome drawbacks. These AHA are often irritants and have a poorbioavailability: they sometimes penetrate too rapidly into the deeplayers of the skin. In Parfums Cosmétiques Arômes 122,66-72 (April, May1995) improvements proposed by the suppliers of raw cosmetic materialsare described to make these compounds less irritant and to slow downtheir penetration into the skin.

A proposed improvement attempt consists of encapsulating these AHA incapsules in order to slow down the diffusion of AHA. Unfortunately, itis difficult to know with exactitude the percentage of activeingredients encapsulated and even more difficult to evaluate thepercentage of active ingredients liberated into the skin.

Another attempt consists of lipophilising these AHA by grafting alipophile compound (fatty alcohol, alkyl chains) by esterification.However, the action of these compounds is reduced because of theirlipophile nature. In fact, they diffuse only with great difficulty intothe stratum corneum into which they are stopped by the presence ofaqueous compartments in the intercorneocytory spaces.

We have expanded this reflection to saturated and unsaturated fattyacids. In fact, the corneal layer is made up of a compact mass of 20layers of inactive cells, embedded in a system of double lamellar layersof lipids. This structure of the stratum corneum as well as thelipophile nature of the lipid barrier protects the skin against thedrying out provoked by the imperceptible loss of transepidermal water.Cosmetic and/or dermatological compositions have, amongst others, avocation of acting on the protection function of the skin and to improvethe appearance. If the intercellular lipids of the corneal layer arealtered, the skin no longer protects.

Unsaturated fatty acids, such as linoleic acid, are an important factorfor the construction and repair of the lipid barrier. They function asprecursor molecules for the synthesis of a signal substance whichcontrols the proliferation and the activity of the cells.

In addition, unsaturated fatty acids also directly take part in theregulation of the cutaneous permeability. These are non occlusivelipophile substances capable of making a more or less continuous film atthe cutaneous surface but above all likely to incorporate themselves inthe intercellular cement thus playing an active role in the regulationof hydration. Their biological activity is regulated by the position ofthe double link closest to the terminal methyl group.

The clinically observed cases of cutaneous alteration like acne showthat a sufficient supply of unsaturated fatty acids in the skin isnecessary to maintain the functioning of the lipid barrier.

Unsaturated fatty acids thus have an essential role in the physiology ofthe skin. Their topical administration however poses problems, problemsthat we propose to resolve by vectoring them.

The action of these compounds are reduced as they only diffuse withdifficulty into the stratum corneum: they are stopped by the presence ofaqueous compartments contained in the intercorneocytary spaces. Thegrafting of these acids onto α-butylglucoside increases the hydrophilenature and thus optimises the penetration of these active ingredientsinto the epidermis.

In addition, the penetration of these compositions into the epidermisposes a problem because of their lipophile nature. Their introductioninto emulsions stabilised by a monolayer of tensioactives practicallydoes not improve this state of fact given that these emulsions break assoon as they are applied onto the skin. An oily phase containing theunsaturated fatty acids thus rests on the surface of the skin. Thanks tothe invention the increase of the hydrophilic nature by the hydroxylfunctions free from the glucose part improves penetration and allows anoptimised usage of unsaturated fatty acids in water in oil or oil inwater emulsions.

By extension, this principle can be applied to numerous active lipophilecompounds with a physiological action on the skin. As an example, we cancite the esterifiable derivatives of the lipophile vitamins A,D,E or F,essential oils, solar filters, anti-inflammatories as well asbio-stimulant agents of lipids and/or protein syntheses. The documentFR-A-94 12005 exposes different solutions consisting of preparing oilemulsions in specific water.

Certain cosmetic, dermatological pharmaceutical active ingredientsand/or food ingredients are unstable as they are sensitive to exteriorfactors like light or heat.

Moreover, different means have been used to stabilise these compounds.One of these means lies for example in blocking the sensitive site byesterification with phosphate, sulphate, and alkyl derivatives and toemploy these derivatives instead of the non-modified compound. Thesederivatives have a less good activity and are sometimes more toxic thanthe active ingredient free by the presence of phosphated, sulphated oralkyl residues.

Another means consists of blocking the site with a glycosidicderivative. A precursor of active ingredients is thus obtained whichafter application on the skin is stopped by the cutaneous enzymes or ishydrolysed after oral administration. The active ingredient is thenliberated. Thus, the patent EP-A-627441 describes the. preparation andthe use of glucosylate of ascorbic acid topically, stopped by theenzymes of the skin which then liberate ascorbic acid. But the use ofsuch derivatives also brings about the liberation of glucose at thesurface of the skin which favours the development of pathogeniccutaneous flora.

The applicant has now found in an unexpected manner that certainalkylmonoglucosides and more particularly α-butylglucoside, allowed allthese improvements whilst avoiding the problems described in the priorart.

The present invention thus has as its principal object the preparationand use of alkylmonglucosides and more particularly α-butylglucoside ina food, cosmetic, dermatological or pharmaceutical composition.

The alkylglucosides, more particularly α-butylglucoside, as well ascertain alkylglucoside esters, can be obtained by enzymatic route as isdescribed in the patent application PCT/FR92/00782 belonging to theapplicant.

The products stemming from this process are anomerically pure (α) andare monoglucosides. Because of the quasi absence of anomer β, thecompounds thus obtained, have specific physico-chemical properties, suchas the point of fusion and the solubility. The amphiphile nature makesthe α-butylglucoside miscible in all proportions with saturated andunsaturated acids in a melted medium and soluble in the aqueous mediumsand polar organic synthesis solvents.

TABLE n°1 solubility of α-butylglucoside in water and in differentsolvents Solubility at 20° C. Solvent % (p/p) Water 60 Ethanol (95/96%)70 Glycerol 70 Acetonitrile 50 Dichloromethane 70 Dioxane 70Polyethylene glycol 200 70 Propylene glycol 70

We can modify the active ingredients and/or ingredients byα-butylglucoside to make them lipophile or hydrophilic. It thus becomespossible to modulate their physico-chemical behaviour as well as theirpenetration, which makes it possible to strongly reduce any risk ofirritation.

The active ingredients possessing at least one acid or ester functionare able to be grafted with the aid of a chemical or enzymatic catalyst,notably hydrolysed trialglycerols (E.C no. 3.1.1.3) which can act ascarboxyesterase. We have used Novozym® or Lipozyme® as they are easilyaccessible commercial enzymes. The active ingredients to which theinvention applies concerns those comprising for example:

At least one acid function and more particularly amino and α-hydroxyacids such as glycolic acid, lactic acid, malic acid, tartaric acid,citric acid, gluconic acid, salicylic acid and serine.

An acid function and notably butyric acid, saturated and unsaturatedfatty acids, and more particularly oleic acid, erucic acid, ricinoleicacid, linoleic acid, and alpha and gamma linoleic acid.

At least one ester function like methyl lactate, ethyl lactate, butyllactate or any other esterified derivative of the acids mentioned above.It can be cetone esters and notably dihydroxyacetone ester. The esterused according to the invention includes one or several ester functionswith linear or branched chain, saturated or unsaturated, with from 2 to25 atoms of carbon, possibly comprising one or several substituents.

At least one carboxylic esterifiable function, and notably, vitaminderivatives such as retinol (vitamin A) and its derivatives (notablyretinoic acid), ascorbic acid (vitamin C) and its derivatives,tocopherols, amongst which are vitamin E and the D vitamins, as well asamino acids, peptides and their derivatives. It can also concernderivatives of polyphenols and more particularly polyhydroxylatedderivatives of flavan and especially flavan-3-ol.

By vectorized active ingredients, we mean the coupling by a chemicalcovalent link of the active ingredient or ingredient to the position 6of an alkylmonoglucoside and more particularly to α-butylglucoside, of aformula

in which R₁ is C₂₋₁₈ alkyl, linear or branched radical, preferably abutyl radical.

The vectorized active ingredients can be used according to the inventionin a quantity from 0.1 to 50% in weight, and preferably from 0.5 to 50%in weight when it concerns vectorized cetone ester and notablyvectorized dihydroxyacetone ester and from 0.1 to 10% in weight whereother vectorized active ingredients are concerned.

As an example, and without it being considered as limiting, thepreparation of two types of derivatives is described according to thepresent invention.

EXAMPLE 1 Preparation of Lactic Ester of α-Butylglucoside

In a flask adaptable to a rotating evaporator, 120 g of α-butylglucosideis placed in 1 litre of butyl L-lactate at 60° C. under vacuum. 100 g ofNovozym®, of the lipase of Candida antarctica immobilised on a solidsupport (Novo Industri, Denmark). The solution is placed again undervacuum (15 mbars) at 60° C. for at least 48 hours.

At the end of the reaction, the reaction medium is centrifuged andfiltered to completely eliminate the Novozym®. The filtrate is collectedfor purification. The lactate of a-butylglucoside is purified by twosuccessive extractions in hexane. The butyl lactate is extracted in theorganic phase. The lactate of α-butylglucoside is collected in theaqueous phase which is decolourised in active carbon. 2.5% (p/v) ofactive carbon is added into a molar solution of lactate ofα-butylglucoside. The whole contents is left to incubate underagitation, for 3 hours at 60° C., then filtered to eliminate the activecarbon.

EXAMPLE 2 Preparation of Lactic Ester of α-Butylglucoside

In a flask adaptable to a rotating evaporator, the α-butylglucoside (100mM) and methyl L-lactate (500 mM) is placed in 20 ml of methyl-2 butanolat 60° C. under vacuum. 100 g/l of Novozym® is added, from the lipase ofCandida antarctica immobilised on a solid support (Novo Industri,Denmark). The solution is placed again at 60° C. under vacuum (200mbars).

The filtrate is collected for purification. The lactate ofα-butylglucoside is purified by two successive stages. The Novozyme® iseliminated by filtration. The methyl lactate and methyl-2-butanol-2 areeliminated by evaporation under vacuum (80° C.-10 mbars).

EXAMPLE 3 Preparation of 6-O-Oleylα-Butylglucoside by EnzymaticEsterification

A medium has been made up with 1.2 g of oleic acid and 1 g ofα-D-butylglucoside. The mixture is brought to 65° C., fusion temperatureof α-butylglucoside. After homogenisation, 1 g of Lipozyme®, from thelipase of Mucor miehei immobilised on a solid support (Novo Industri,Denmark) is added.

After 3 days of incubation, the reaction medium is diluted with 20 ml ofdiethyl ether, then filtered in order to eliminate the Lipozyme®. Afterfiltration, 50 ml of soda (0.02N) is added, in order to make theresidual oleic acid in the form of a sodium salt, as well as theresidual α-butylglucoside soluble in the aqueous phase.

The organic phase containing the oleic acid ester is evaporated undervacuum in order to eliminate any trace of residual solvent.

As a variant, Novozyme®, at a concentration of 5% in weight in relationto the substrates, has been substituted for Lipozyme®. Equally aninferior alkyl (methyl or ethyl, for example) oleate can be substitutedfor oleic acid.

Measures by HPLC have shown that more than 95% of the acid can beconverted whatever the enzyme and the acylated compound chosen. Howeverthe use of Novozym® as biocatalyst leads to higher conversion with morethan 98% of the two substrates which have been converted.

The control of the reaction by Thin Layer Chromatography (TLC) has showna formation of a single product when the Novozyme® has been used asbiocatalyst.

The structure of the products has been determined by Mass Spectrometry(MS) Nuclear Magnetic Resonance (NMR).

After acylation of α-butylglucoside with the oleic acid with Novozym® asa catalyst, the product has been purified.

The results of MS show a molecular ion at m/z=523.3 [M+Na]⁺, Mcorresponds exactly to the molecular mass of monooleate ofα-butylglucoside. The results of NMR have demonstrated thatα-butylglucoside has been grafted exclusively on position 6 of themolecule.

The chemical displacement of the proton at 5.42 ppm confirms thepresence of a carbon-carbon unsaturation coming from the oleic part ofthe molecule 6-O-oleyl α-butylglucoside. The iodine indices from thereaction medium at the beginning (mixture of oleic acid andα-butylglucoside) and at the end (6-O-oleyl α-butylglucoside) of theenzymatic esterification are the same at less than 3% of error (43.7 and42.7 respectively). That shows that no oxydation of the double linkoccurred during the process of enzymatic esterification.

EXAMPLE 4 Synthesis of a Mixture of α-Butylglucoside Esters

The acylation of α-butylglucoside is proceeded to by a commercialmixture of linoleic acid (60.5% molar), oleic acid (32.7% molar) andlinoleic acid (6.8% molar) in the absence of solvent, at 60° C. under areduced pressure of 20 mbars, with an initial acid/ α-butylglucosidemolecular yield of 1, and in the presence of Novozym® to an initialconcentration of 5% in weight in comparison to the total weight of theacid reactives and α-butylglucoside.

The water generated by the reaction has been evaporated underestablished reduced pressure, in order to displace the thermodynamicequilibrium in favour of the synthesis of esters.

The esters produced have been collected by adding a bit of hexane thenby eliminating the enzyme by filtration.

The conversion of the initial acids into esters of α-butylglucoside hasbeen almost total (>95%).

As an example, and without it being considered as limiting, theimprovement of cutaneous and ocular irritation created by the graftingof a preparation onto α-butylglucoside is described below.

EXAMPLE 5 Comparison of the Cytotoxicity of Lactic Acid and of Lactateof α-Butylglucoside

We have quantified and compared in vitro, the cytotoxicity of lacticacid and of lactate of α-butylglucoside on a skin model. This skin modelis made up of a matrix of collagen on which rests an epidermisreconstituted from keratinocytes and presenting a similar architectureto that of the human epidermis. The transformation of tetrazolium salt(MTT) in blue crystals of Formozan is proportional to the activity ofthe dehydrogenase succinate, (a mitochondrial enzyme). As a consequence,the more the epidermis contains living cells, the larger thetransformation by the dehydrogenase succinate of the MTT in blueFormozan crystals will be. The quantity of Formozan is measured in thespectrophotometer. The cellular viability, carried out to determine thetoxicity of the product, is calculated according to the formula:

% viability=OD produced/OD cellular control×100

The cellular control is carried out without issue. A referencetensioactive SDS (Sodium Dodecyl Sulphate) at 1.5 mg/ml is equallytested as a cytotoxic control.

If the cellular viability is between 70 and 50%, the products areconsidered as slightly cytotoxic.

If it is lower than 50%, the products are considered as cytotoxic.

Table 2 shows the results of the evaluation.

TABLE n°2 MTT dosage on skin model optical densities obtained at 540 nmand percentage of viability of epidermic cells in comparison to thecontrol Lactate of α- SDS at Lactic acid butyl glucoside Trial Control1-5 mg/ml at 5% at 5% OD trial 1 0.324 0.046 0.044 0.242 OD trial 20.363 0.056 0.066 0.265 Average 0.343 0.051 0.055 0.255 % viability in100% 14.8% 16% 74.5% comparison to the control

The lactate of α-butylglucoside, vectorized lactic acid, is lessirritant than lactic acid. Lactate of α-butylglucoside at 5% is notcytotoxic.

EXAMPLE 6 Comparison of the Ocular Toxicity of lactic Acid and ofLactate of α-Butylglucoside (Vectorized Lactic Acid)

The BCOP test (bovine corneal opacity and permeability) makes itpossible to evaluate the ocular irritation indice in vitro on corneasfrom abattoir beef carcasses. It measures two parameter of ocularirritation: the opacity and cellular permeability.

The opacity is defined as the difference of light transmission between atreated cornea and a control cornea. The permeability is equal to theoptical density (OD) measured in a spectrophotometer at 490 nm.

The irritation score in vitro is established by using the followingformula:

Score in vitro=opacity value+1.5 times the value of OD.

The objective values are combined and the scores of ocular irritation invitro are compared to an irritation scale previously established. Ingeneral, the irritant potential is classed in three categories weak (0to 25), moderate (25.1 to 55) and severe (55.1 and +). The ocularirritation indice in vitro of lactate of α-butylglucoside at 36% is zeroalthough that of lactic acid at 5% and 10% are, respectively from 1.93and from 41.11.

Our preparation of α-butylglucoside at 36% thus belongs to the categoryof non-irritant products, whilst lactic acid at 10% belongs to thecategory of moderately irritant products.

What is claimed is:
 1. A method of causing enhanced transcutaneous ortransmucous permeation in a patient of an active ingredient in apharmaceutical, dermatological, cosmetic or food composition,comprising: applying said composition to skin or mucous membrane of saidpatient, said composition comprising said active ingredient as avectorized compound of the formula:

wherein R₂ is a —CO—R group in which R is a

radical, with X being an aliphatic chain, and R₁ is linear or branchedC₂-C₁₈ alkyl.